The translation of cellular messenger RNAs into distinct structural and functional proteins is central to gene expression in all domains of life an serves as a critical conduit for proteome capacity and diversity. The multistep and highly regulated process of translation is carried out by the ribosome, a two-subunit, RNA-protein assembly that is composed of 70 distinct gene products in bacteria and more than 80 distinct gene products in humans. Although the structural and functional features of the translation mechanism are thought to be conserved throughout evolution, the therapeutic administration of small-molecule antibiotics targeting the bacterial translation machinery continues to serve as a critical safeguard against existing and emerging infectious diseases around the world. Hence, key aspects of the translation mechanism and/or ribosome structure in humans must be distinct from bacteria. The loss of translation control in human cells is linked to cancerous growth and a growing body of knowledge suggests that ribosomes within cancer cells may be physically and functionally distinct. Correspondingly, knowledge of the common and distinct features of bacterial and human ribosomes offers the potential to enable improvements in the efficacies of existing antibiotics, the development of potentially novel means for antibiotic intervention and holds the promise of translation-specific strategies for cancer treatment. Progress on these fronts requires quantitative descriptions of structure-function relationships in the translation machinery of bacteria and humans as well as dynamic aspects of the translation mechanism, including time-dependent changes in ribosome composition and conformation that occur during processive protein synthesis reactions. The proposed research aims to elucidate the molecular basis of ribosome function, the origins of translational fidelity and the molecular mechanisms of antibiotic action using a battery of state-of-the-art biophysical approaches. The methods include single-molecule Total Internal Reflection Fluorescence imaging and rapid-stopped flow kinetics measurements, together with collaborative and complementary efforts in molecular dynamics simulation and high-resolution structure determination. In so doing, we aim to delineate quantitative kinetic and structural models of bacterial and human ribosome function to define key distinctions that will inform on new opportunities for small-molecule interventions for the treatment of infectious pathogens and human disease.